Rho controls cortical F-actin disassembly in addition to, but independently of, secretion in mast cells

Localized disassembly of cortical F-actin has long been considered necessar y for facilitation of exocytosis. Exposure of permeabilized mast cells to c alcium/ATP induces cortical F-actin disassembly (calmodulin-dependent) and secretion (calmodulin-independent). The delay in the onset of secretion is characteristic for the calcium/ATP response and is abolished by GTP. Here w e report that a constitutively active mutant of Rho (V14RhoA) enhanced both secretion and cortical F-actin disassembly. In addition, V14RhoA mimicked GTP by abolishing the delay in secretion. Inhibition of Rho by C3 transfera se prevented both secretion (similar to 80%) and F-actin disassembly (simil ar to 20%). Thus, both Rho GTPase and calcium/calmodulin contribute to the control of cortical F-actin disassembly. Stabilization of actin filaments b y high concentrations of phalloidin or by a calmodulin-inhibitory peptide ( based on the calmodulin-binding domain of myosin light chain kinase) did no t affect the extent of secretion or the secretion-enhancing effects of V14R hoA These results further support the existence of divergent, Rho-dependent , pathways regulating actin and exocytosis. Furthermore, compound Y-27632, a specific inhibitor of Rho-associated protein kinase (p160(ROCK)), attenua ted the Rho-induced loss of cortical F-actin without affecting secretion. A model is presented in which Rho regulates secretion and cortical F-actin i n a manner dependent on and/or synergistic with calcium.